BACKGROUND: Evidence indicates that senescence can affect essential dental pulp functions, such as defense capacity and repair, consequently affecting the successes of conservative endodontic treatments. This study aims to evaluate the effects of senescence on the morphology, migration, proliferation, and immune response of human dental pulp cells.
METHODS: Cells were treated with doxorubicin to induce senescence, confirmed by β-galactosidase staining. Morphological changes, cellular proliferation, and migration were evaluated by scanning electron microscopy, trypan blue cells, and the scratch method, respectively. Modifications in the immune response were evaluated by measuring the genes for pro-inflammatory cytokines tumor necrosis factor alpha and interleukin (IL)-6 and anti-inflammatory cytokines transforming growth factor beta 1 and IL-10 using the real time polymerase chain reaction assay.
RESULTS: An increase in cell size and a decrease in the number of extensions were observed in senescent cells. A reduction in the proliferative and migratory capacity was also found in senescent cells. In addition, there was an increase in the gene expression of inflammatory cytokines tumor necrosis factor alpha and IL-6 and a decrease in the gene expression of IL-10 and transforming growth factor beta-1, suggesting an exacerbated inflammatory situation associated with immunosuppression.
CONCLUSIONS: Cellular senescence is possibly a condition that affects prognoses of conservative endodontic treatments, as it affects primordial cellular functions related to this treatment.

Dental pulp under physiological conditions has a defense function, repair capacity, and important mechanisms in pathological processes. In addition, the dental papilla is involved in important defense processes and an essential function in the pulp revascularization process. It is known that dental pulp and apical papilla undergo a natural aging process, in addition to stressful situations such as bruxism, inflammation, and infections. Both aging and stressful situations can lead to cellular senescence. Some evidence indicates that the changes resulting from this cellular state can directly affect the efficiency of cells in these tissues and affect conservative and regenerative clinical treatments. Thus, it is necessary to understand the causes and consequences of cellular senescence in addition to the development of methods for senescence prevention. This review aims to provide an overview of possible causes and consequences of senescence in dental pulp and stem cells from apical papilla and discusses possible methods to prevent this cellular state.

OBJECTIVE: To assess in vitro the effect of two novel phase separated borosilicate glasses (PSBS) in the system SiO2 -B2 O3 -K2 O-CaO-Al2 O3 on dental pulp cells; and to compare their bioactivity and mechanical properties to a conventional fluoroaluminosilicate glass ionomer cement namely FUJI IX.
METHODS: The cytocompatibility assessment of the two novel borosilicate glasses, one without alumina (PSBS8) and one containing alumina (PSBS16), was performed on cultured primary human pulp cells. Alamar blue assay was used to assess cell metabolic activity and cell morphology was evaluated by confocal imaging. The bioactivity in Stimulated Body Fluid was also evaluated after 1 and 3 weeks of immersion using SEM-EDX analysis. Vickers microhardness and flexural strength were assessed after incorporating the glass particles into a commercial glass ionomer cement (GIC) liquid containing both polyacrylic and polybasic carboxylic acid.
RESULTS: The data revealed that the two borosilicate glasses enhanced cell viability ratios at all-time points in both direct and indirect contact assays. After 3 days of contact, PSBS8 without alumina showed higher viability rate (152%) compared to the PSBS16 containing alumina (145%) and the conventional glass ionomer particles (117%). EDX analysis confirmed an initial Ca/P ratio of 2.1 for 45S5K and 2.08 for PSBS8 without alumina after 3 weeks of immersion. The cement prepared using PSBS8 showed significantly higher Vickers hardness values (p = .001) than that prepared using PSBS16 (46.6 vs. 36.7 MPa). After 24 h of maturation, PSBS8 (without alumina) exhibited a flexural strength of 12.9 MPa compared to a value of 16.4 MPa for the commercial control. PSBS8 without alumina had a higher strength than PSBS16 with alumina, after 1 and 7 days of maturation (p = .001).
CONCLUSIONS: The present in vitro results demonstrated that the borosilicate bioactive glass without alumina enhanced pulp cell viability, spreading and acellular bioactivity better than the conventional GIC and the experimental borosilicate glass containing alumina.

Dental pulp cells interact with immunogenic components such as LPS (lipopolysaccharide) or LTA (lipoteichoic acid) released from microorganisms in carious lesions. In the present investigation, the formation of the pro-inflammatory cytokines TNFα and IL-6 in LPS- or LTA-stimulated cells from the dental pulp interface and pulp fibroblasts was analyzed in the presence of the resin monomer 2-hydroxyethyl methacrylate (HEMA) under varying cellular redox conditions.
Human pulp fibroblasts (HPC) or cells from the dental pulp interface expressing an odontoblast phenotype (hOD-1) were exposed to LTA, LPS or HEMA for 1 h or 24 h. Redox homeostasis was modified by the prooxidant BSO (L-buthionine sulfoximine) or the antioxidant NAC (N-acetyl cysteine). Formation of TNFα or IL-6 was analyzed by ELISA, and cell survival was determined by a crystal violet assay. Statistical analyses were performed using the Mann-Whitney-U-test.
Secretion of TNFα was not detected in LPS- or LTA-stimulated HPC or hOD-1, and IL-6 was not found after a short exposure (1 h). After a 24 h exposure, LPS induced a 3-fold increase in IL-6 formation in HPC, while LTA stimulated IL-6 release about 20-fold. Likewise, LTA was more effective than LPS in hOD-1 stimulating IL-6 levels about 50-fold. HEMA inhibited the LPS- and LTA-induced IL-6 release, and this effect was enhanced by BSO but counteracted by NAC in both cell types. IL-6 release was independent of cell survival rates.
The protective immune response in odontoblasts and pulp fibroblasts is impaired by monomers such as HEMA through the disturbance of the redox homeostasis.

Lipopolysaccharide (LPS) is widely used for induction of inflammation in various human tissues, including dental pulp. The purpose of this study was to summarize current medical literature focusing on (1) cell types used by researchers to simulate dental pulp inflammation, (2) LPS variants utilized in experimental settings and how these choices affect the findings. Our study was conducted in accordance with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). We searched for studies reporting outcomes of lipopolysaccharide application on dental pulp cells in vitro using electronic databases: MEDLINE, Web of Science and Scopus. Having gathered data from 115 papers, we aimed to present all known effects LPS has on different cell types present in dental pulp. We focused on specific receptors and particles that are involved in molecular pathways. Our review provides an essential foundation for further research using in vitro models of pulpitis.

The cellular atlas of the stroma is not well understood. Here, the cell populations in human dental pulp through single-cell RNA sequencing are profiled. Dental pulp stem cells, pulp cells, T cells, macrophages, endothelial cells, and glial cells are identified in human dental pulp. These cells support each other through sending growth signals. Based on the appearance of ligand-receptor pairs between two cell populations, pulp cells have the greatest communication with other cell types, while T cells have the least communication. In addition, T cells expressing TLR1, TLR2, and TLR4, and endothelial cells expressing TLR4, monitor bacterial invasion. These findings provide the census of normal dental pulp.

Healthy pulp tissue plays an important role in normal function and long-term retention of teeth. Mesoporous bioactive glass (MBG) as a kind of regenerative biomaterials shows the potential in preserving the vital pulp. In this study, MBG prepared by organic template method combined with sol-gel method were used in human dental pulp cell culture and ectopic mineralization experiment. Real-Time PCR was used to explore its ability to induce odontogenic differentiation of dental pulp cells. MBG and rat crowns were implanted under the skin of nude mice for 4 weeks to observe the formation of pulp dentin complex. We found that MBG can release Si and Ca ions and has a strong mineralization activity in vitro. The co-culture of MBG with human dental pulp cells promoted the expression of DMP-1 (dentin matrix protein-1) and ALP (alkalinephosphatase) without affecting cell proliferation. After 4 weeks of subcutaneous implantation in nude mice, the formation of hard tissue with regular structure under the material could be seen, and the structure was similar to dentin tubules. These results indicate that MBG can induce the differentiation of dental pulp cells and the formation of dental pulp-dentin complex and has the potential to promote the repair and regeneration of dental pulp injuries.

The purpose of this study is to analyze the influence of InGaAlP diode laser (660 nm) with or without an odontogenic medium (OM) in the functional activity of OD-21 cells. Undifferentiated OD-21 pulp cells were cultivated with or without OM and divided into four groups (n = 5): nonirradiated control (C -), nonirradiated + OM (C +), irradiated (L -), and irradiated + OM (L +). Laser application was performed in two sessions of a 24-h interval with an irradiance of 11.3 mW/cm2, energy density of 1 J/cm2, and total cumulative energy/well of 4.6 J. Cell proliferation, VEGF-164 expression, mineralization, and expression of Alp, Runx2, and Dmp1 genes, as well as immunolocalization of RUNX2 and MEPE proteins, were evaluated. Data were analyzed by statistical tests (α = 0.05). All studied groups showed a similar increase in cell proliferation with or without OM. After 7 and 10 days, a significatively higher concentration of VEGF-164 in L - group when compared to C - group was observed. A significant increase in mineralized nodules in the L + was noted when compared to C + in the same conditions. Photobiomodulation upregulated significantly Runx2 and Dmp1 expression after 10 days in L - and after 7 days in L + , with downregulation of Dmp1 after 10 days in L + group. Immunolocalization of RUNX2 and MEPE was expressive after 7 days of culture in the cytoplasm adjacent to the nucleus with a decrease after 10 days, regardless of the presence of OM. Photobiomodulation enhances metabolism associated with angiogenesis, gene expression, and mineralization regardless of the odontogenic medium in OD-21 cells.

The effects of surface pre-reacted glass-ionomer (S-PRG) filler on pulpal cells and on the composition of dentinal deposits were investigated. Proliferation (CCK-8), cytotoxicity (LDH), and differentiation activity (ALP) tests, along with cell morphology observations, were conducted at 6 and 24 h after treatment of pulpal cells with different S-PRG filler eluate concentrations. Dentinal surfaces were immersed in deionized water or S-PRG filler eluate followed by immersion in deionized water or simulated body fluid and observed under scanning electron microscope and elemental analysis using energy dispersive x-ray spectrometer. At 24 h, there were significant differences in CCK-8 and ALP activity values between the groups in a concentration-dependent manner. LDH test data were not significantly different among the groups. Cell morphology was not altered at either exposure time. However, decreased cellular density was observed with the highest eluate concentration. Crystalline deposits and occluded dentinal tubules were observed in samples immersed in S-PRG filler with a later immersion in simulated body fluid, which also showed higher concentrations of certain ions compared to surfaces that were not initially treated with S-PRG filler. The lowest two eluate concentrations did not show significant toxicity. S-PRG enhanced the effect of simulated body fluid in the formation of mineral deposits.

Pulpal inflammation is known to be mediated by multiple signaling pathways. However, whether melatonin plays regulatory roles in pulpal inflammation remains unclear. This study aimed at elucidating an in situ expression of melatonin and its receptors in human pulpal tissues, and the contribution of melatonin on the antagonism of lipopolysaccharide (LPS)-infected pulpal fibroblasts.
Melatonin expression in pulpal tissues harvested from healthy teeth was investigated by immunohistochemical staining. Its receptors, melatonin receptor 1 (MT1) and melatonin receptor 2 (MT2), were also immunostained in pulpal tissues isolated from healthy teeth and inflamed teeth diagnosed with irreversible pulpitis. Morphometric analysis was subsequently performed. After LPS infection of cultured pulpal fibroblasts, cyclo-oxygenase (COX) and interleukin-1 β (IL-1 β) transcripts were examined by using reverse transcription-polymerase chain reaction (RT-PCR). Analysis of mRNA expression was performed to investigate an antagonism of LPS stimulation by melatonin via COX and IL-1 β induction. Mann-Whitney U test and One-way ANOVA were used for statistical analysis to determine a significance level.
Melatonin was expressed in healthy pulpal tissue within the odontoblastic zone, cell-rich zone, and in the pulpal connective tissue. Furthermore, in health, strong MT1 and MT2 expression was distributed similarly in all 3 pulpal zones. In contrast, during disease, expression of MT2 was reduced in inflamed pulpal tissues (P-value< 0.001), but not MT1 (P-value = 0.559). Co-culturing of melatonin with LPS resulted in the reduction of COX-2 and IL-1 β expression in primary pulpal fibroblasts, indicating that melatonin may play an antagonistic role to LPS infection in pulpal fibroblasts.
Human dental pulp abundantly expressed melatonin and its receptors MT1 and MT2 in the odontoblastic layers and pulpal connective tissue layers. Melatonin exerted antagonistic activity against LPS-mediated COX-2 and IL-1 β induction in pulpal fibroblasts, suggesting its therapeutic potential for pulpal inflammation and a possible role of pulpal melatonin in an immunomodulation via functional melatonin receptors expressed in dental pulp.